Dry composition comprising an extrusion enhancer

ABSTRACT

Disclosed is a dry composition comprising one or more polyols, which upon addition of an aqueous medium forms a substantially homogenous paste suitable for use in haemostasis procedures. The paste reconstitutes spontaneously upon addition of the liquid; hence no mechanical mixing is required for said paste to form. The composition may further comprise an extrusion enhancer, such as albumin. Also disclosed are methods of preparing said dry composition, a paste obtained from said dry composition and uses of said dry composition or paste for medical and surgical purposes.

RELATED APPLICATIONS

This application is a divisional of U.S. application Ser. No.15/102,994, filed Jun. 9, 2016, which is the U.S. National Stage ofInternational Application No. PCT/DK2014/050421, filed on Dec. 10, 2014,published in English, which claims priority under 35 U.S.C. § 119 or 365to Denmark Application No. 2013 70758, filed Dec. 11, 2013. The entireteachings of the above applications are incorporated herein byreference.

TECHNICAL FIELD

The present disclosure relates to a dry composition suitable for use inhaemostasis and/or wound healing, wherein the dry composition comprisesone or more polyols and forms a paste spontaneously upon addition of anaqueous medium. The composition further comprises an extrusion enhancer,such as albumin. The disclosure further relates to methods of preparingsaid dry composition and use of said composition.

BACKGROUND

Protein-based haemostatic materials such as collagen and gelatine arecommercially available in solid sponge and loose or unpacked powder formfor use in surgical procedures. Mixing of the loose or unpacked powderwith a fluid such as saline or thrombin may form a paste or slurry thatis useful as a haemostatic composition for use in cases of diffusebleeding, particularly from uneven surfaces or hard to reach areas,depending on mixing conditions and relative ratios of the materials.

Conventional haemostatic pastes are prepared at the point of use bymechanical agitation and mixing of loose powder and liquid to provideuniformity of the composition. Mixing of the powder and fluid may beconducted in a container, such as a beaker. Such mixing requirestransfer of the powder from its original container to the beaker,addition of the fluid to the beaker containing the powder, and thenkneading of the mixture to form the paste. Only after the paste is thusformed may the paste be placed into a delivery means or applicator, e.g.a syringe, and applied to the wound.

WO 03/055531 relates to a container comprising a fixed amount ofhaemostatic agent in powder form, such as gelatine powder. Upon additionof a suitable amount of liquid, mechanical mixing within the containeris performed by closing the lid and shaking the container. The resultantputty-like haemostatic paste can then be removed from the container andapplied to a patient to promote haemostasis.

Alternately, attempts have been made to preload one syringe (Syringe I)with loose gelatine powder, and a second syringe (Syringe II) withliquid. When it is time to make a paste, Syringes I and II are connectedvia a luer lock and the solution in Syringe II is pushed into Syringe I.By attempting to pass the solution and powder repeatedly back and forthbetween Syringes I and II, a homogeneous paste may or may not be formed.Often in a surgical situation, a haemostatic paste with optimalpowder:liquid ratio cannot be obtained due to insufficient mixing of thepowder and the liquid in a syringe. Even if such methods of mixing aresuccessful in forming a paste, the time and mechanical effort requiredto form the paste are undesirable or even unacceptable. Also the mixingcan affect the final density of the paste (too intense mixing may resultin a lower density paste) and hence inconsistent consistency of thepaste from time to time.

Floseal Haemostatic Matrix (Baxter) is a kit for producing a haemostaticgelatine paste. The gelatine paste is produced by first making athrombin solution and then transferring the gelatin matrix-thrombinsolution mixture back and forth between two connected syringes for atotal of at least twenty passes. The paste can then be applied to apatient to promote haemostasis directly from the syringe.

Likewise, Surgiflo® Haemostatic Matrix (Ethicon) is a kit for producinga haemostatic gelatine paste comprising thrombin, which is prepared bytransferring the gelatin matrix-thrombin solution mixture back and forthbetween two connected syringes for a total of at least 6 passes.

US 2005/0284809 relates to a method for preparing a haemostatic pastethat more readily absorbs aqueous liquids, such that less mechanicalforce and time is required in order to form a flowable haemostaticpaste. The paste of US 2005/0284809 is prepared from compressedhaemostatic powder particles and to prepare the paste, it must betransferred back and forth between connected syringes for a total of atleast 5 passes.

WO 2011/151400 relates to a process for making a dry haemostaticcomposition comprising a coagulation inducing agent such as thrombin anda biocompatible polymer such as gelatine. The coagulation inducing agentand the biocompatible polymer are mixed to form a paste and the paste issubjected to lyophilisation. The resulting dry composition isreconstituted by transferring the composition and a diluent back andforth between two connected syringes for a total of at least twentypasses as described previously.

Mixing procedures and manipulations are time consuming and maypotentially compromise the sterility of the haemostatic paste. It wouldbe desirable if a haemostatic composition could be provided which wouldeliminate the need for such undesirable mixing requirements.

WO 2013/185776 discloses a dry paste composition suitable for woundhealing and haemostatic use which reconstitutes spontaneously to form aflowable paste, i.e. without any mixing required, upon addition of anaqueous medium. The dry composition is prepared by mixing a crosslinkedbiocompatible polymer, one or more polyols and an aqueous medium toprepare a paste and then lyophilising the paste to obtain the drycomposition.

WO 2013/060770 discloses a gelatine paste composition, wherein saidcomposition comprises an extrusion enhancer. It is shown that theprovision of extrusion enhancers, such as albumin, in the appropriateamount, enables the use of higher gelatine concentrations which may inturn improve the haemostatic properties of such products. WO 2013/060770is incorporated by reference in its entirety.

It would be desirable to provide spontaneously reconstituting dry pastecompositions which are easier to extrude from a syringe thanconventional flowable paste products. It would also be desirable toprovide spontaneously reconstituting dry paste compositions having ahigher concentration of polymer, e.g. gelatine, than conventionalflowable pastes, with a retained ability to be extruded easily from asyringe.

SUMMARY

The present disclosure relates to an improved dry composition, whichupon addition of an adequate amount of an aqueous medium forms asubstantially homogenous paste suitable for use in haemostasis and woundhealing procedures. The paste forms spontaneously upon addition of theliquid, i.e. no mechanical mixing is required for said paste to form.

The disclosure further relates to a method of preparing said drycomposition comprising the steps of:

-   -   a. providing a biocompatible polymer in powder form, one or more        polyols, an extrusion enhancer and an aqueous medium,    -   b. mixing the biocompatible polymer, the one or more polyols,        the extrusion enhancer and the aqueous medium to obtain a paste,        and    -   c. drying the paste.

The biocompatible polymer is preferably suitable for use in haemostasisand/or wound healing.

Preferably, the extrusion enhancer is albumin, more preferred humanserum albumin.

Uses of the paste formed from the dry composition are likewise coveredby the present disclosure.

DESCRIPTION OF DRAWINGS

FIG. 1. Phase diagram of water. The phase diagram shows, inpressure-temperature space, the lines of equilibrium or phase boundariesbetween the three phases of solid, liquid, and gas.

FIG. 2. Average reconstitution time+/−standard deviation of thefreeze-dried gelatine pastes comprising different polyols of example 6.

DEFINITIONS

A “bioactive agent” is any agent, drug, compound, composition of matteror mixture which provides some pharmacologic, often beneficial, effectthat can be demonstrated in vivo or in vitro. An agent is thusconsidered bioactive if it has interaction with or effect on a celltissue in the human or animal body. As used herein, this term furtherincludes any physiologically or pharmacologically active substance thatproduces a localized or systemic effect in an individual. Bioactiveagents may be a protein, such as an enzyme. Further examples ofbioactive agents include, but are not limited to, agents comprising orconsisting of an oligosaccharide, a polysaccharide, an optionallyglycosylated peptide, an optionally glycosylated polypeptide, anoligonucleotide, a polynucleotide, a lipid, a fatty acid, a fatty acidester and secondary metabolites. It may be used either prophylactically,therapeutically, in connection with treatment of an individual, such asa human or any other animal.

“Biocompatible” refers to a material's ability to perform its intendedfunction without eliciting any undesirable local or systemic effects inthe host.

“Biologically absorbable” is a term which in the present context is usedto describe that the materials of which the said powder are made can bedegraded in the body to smaller molecules having a size which allowsthem to be transported into the blood stream. By said degradation andabsorption the said powder materials will gradually be removed from thesite of application. For example, gelatine can be degraded byproteolytic tissue enzymes to absorbable smaller molecules, whereby thegelatine when applied in tissues typically is absorbed within about 4-6weeks and when applied on bleeding surfaces and mucous membranestypically within 3-5 days.

“Extrusion enhancer” according to the present disclosure covers anybiocompatible compound capable of facilitating extrusion of a paste froma syringe.

A “gel” is a solid, jelly-like material that can have properties rangingfrom soft and weak to hard and tough. Gels are defined as asubstantially dilute cross-linked system, which exhibits no flow when inthe steady-state. By weight, gels are mostly liquid, yet they behavelike solids due to a three-dimensional cross-linked network within theliquid. It is the crosslinks within the fluid that give a gel itsstructure (hardness) and contribute to stickiness (tack). In this waygels are a dispersion of molecules of a liquid within a solid in whichthe solid is the continuous phase and the liquid is the discontinuousphase. A gel is not a paste or slurry.

“Haemostasis” is a process which causes bleeding to diminish or stop.Haemostasis occurs when blood is present outside of the body or bloodvessels and is the instinctive response for the body to stop bleedingand loss of blood. During haemostasis three steps occur in a rapidsequence. Vascular spasm is the first response as the blood vesselsconstrict to allow less blood to be lost. In the second step, plateletplug formation, platelets stick together to form a temporary seal tocover the break in the vessel wall. The third and last step is calledcoagulation or blood clotting. Coagulation reinforces the platelet plugwith fibrin threads that act as a “molecular glue”.

A “haemostatic agent” according to the present disclosure is abiologically absorbable material. Examples of suitable biologicallyabsorbable materials include but are not limited to gelatine, collagen,chitin, chitosan, alginate, cellulose, polyglycolic acid, polyaceticacid and mixtures thereof.

“International Unit (IU)”. In pharmacology, the International Unit is aunit of measurement for the amount of a substance, based on biologicalactivity or effect. It is abbreviated as IU, UI, or as IE. It is used toquantify vitamins, hormones, some medications, vaccines, blood products,and similar biologically active substances.

A “paste” according to the present disclosure has a malleable,putty-like consistency, such as toothpaste. A paste is a thick fluidmixture of pulverized solid/solid in powder form with a liquid. A pasteis a substance that behaves as a solid until a sufficiently large loador stress is applied, at which point it flows like a fluid, i.e. a pasteis flowable. Pastes typically consist of a suspension of granularmaterial in a background fluid. The individual grains are jammedtogether like sand on a beach, forming a disordered, glassy or amorphousstructure, and giving pastes their solid-like character. It is this“jamming together” that gives pastes some of their most unusualproperties; this causes paste to demonstrate properties of fragilematter. A paste is not a gel/jelly. A “slurry” is a fluid mixture of apowdered/pulverized solid with a liquid (usually water). Slurries behavein some ways like thick fluids, flowing under gravity and being capableof being pumped if not too thick. A slurry may be regarded as a thinpaste, i.e. a slurry generally contains more water than a paste.

“Percentage”. If nothing else in indicated, the percentage is w/w.

“Spontaneous”. The term “spontaneous” is used to describe phenomenaarising from internal forces or causes, which are independent ofexternal agencies or stimuli and which happen within a short period oftime, i.e. preferably within less than about 30 seconds, more preferredwithin less than about 20 seconds, even more preferred within less thanabout 10 seconds or within less than about 5 seconds.

DETAILED DESCRIPTION

The present disclosure relates to a dry composition, which upon additionof an adequate amount of an aqueous medium forms a substantiallyhomogenous paste suitable for use in haemostasis procedures. The pasteforms spontaneously upon addition of the liquid component, i.e. nomechanical mixing is required for said paste to form.

The dry composition may be prepared by a method comprising thesequential steps of:

-   -   a. providing a biocompatible polymer in powder form, one or more        polyols, an extrusion enhancer and an aqueous medium,    -   b. mixing the biocompatible polymer, the one or more polyols,        the extrusion enhancer and the aqueous medium to obtain a paste,        and    -   c. drying the paste.

The present disclosure further relates to a paste suitable for use inhaemostasis and/or wound healing procedures prepared by adding anaqueous medium to the dry composition and use of said paste forpromoting haemostasis and/or wound healing.

Disclosed is also use of the dry composition as described herein in thepreparation of a paste for use in haemostatic and wound healingprocedures.

The advantages of the dry composition disclosed herein are numerous andinclude:

-   -   Less time spent preparing the paste, e.g. bleeding can be        stopped faster.    -   Decreased risk of compromising the sterility of the paste during        preparation due to less handling steps.    -   Decreased risk of making mistakes during preparation due to the        simplified preparation of the paste.    -   Optimal consistency of paste obtained every time.    -   Facile extrusion of paste (less force needed to expel the        haemostatic paste from e.g. a syringe) and thus increased        accuracy of application of the haemostatic paste.    -   Bioactive agents, which are unstable in solution may be added to        the paste prior to drying and will thus be present in the dry        composition of the disclosure. For example, thrombin may be        added to the paste prior to drying, thereby avoiding the        time-consuming and error-prone thrombin dilution steps.

All of the above factors lead to increased patient safety.

Biocompatible Polymer

The present disclosure relates to a biocompatible agent in powder form,which is used to create a paste. The paste is then dried to obtain a drycomposition suitable for use in haemostasis and wound healingprocedures.

The biocompatible polymer of the present disclosure may be a biologic ora non-biologic polymer. Suitable biologic polymers include proteins,such as gelatin, soluble collagen, albumin, hemoglobin, casein,fibrinogen, fibrin, fibronectin, elastin, keratin, and laminin; orderivatives or combinations thereof. Particularly preferred is the useof gelatin or soluble non-fibrillar collagen, more preferably gelatin.Other suitable biologic polymers include polysaccharides, such asglycosaminoglycans, starch derivatives, xylan, cellulose derivatives,hemicellulose derivatives, agarose, alginate, and chitosan; orderivatives or combinations thereof. Suitable non-biologic polymers willbe selected to be degradable by either of two mechanisms, i.e. (1) breakdown of the polymeric backbone or (2) degradation of side chains whichresult in aqueous solubility. Exemplary nonbiologic polymers includesynthetics, such as polyacrylates, polymethacrylates, polyacrylamides,polyvinyl resins, polylactide-glycolides, polycaprolactones, andpolyoxyethylenes; or derivatives or combinations thereof. Alsocombinations of different kinds of polymers are possible.

The paste of the present disclosure may either comprise a singlebiocompatible polymer or a mixture of two or more biocompatiblepolymers.

In one embodiment, the biocompatible polymer is biologically absorbable.Examples of suitable biologically absorbable materials include gelatine,collagen, chitin, chitosan, alginate, cellulose, oxidised cellulose,polyglycolic acid, polyacetic acid and combinations thereof. It will beunderstood that various forms thereof, such as linear or cross-linkedforms, salts, esters and the like are also contemplated for the presentdisclosure.

In a preferred embodiment of the disclosure, the biologically absorbablematerial is gelatine. Gelatine is preferred since gelatine is highlybiologically absorbable. Furthermore, gelatine is highly biocompatible,meaning that it is non-toxic to an animal, such as a human being,when/if entering the blood stream or being in long-term contact withhuman tissues.

The gelatine typically originates from a porcine source, but mayoriginate from other animal sources, such as from bovine or fishsources. The gelatine may also be synthetically made, i.e. made byrecombinant means.

In a preferred embodiment the polymer is cross-linked.

Any suitable cross-linking methods known to a person of skill may beused including both chemical and physical cross-linking methods.

In one embodiment of the present disclosure the polymer has beencross-linked by physical means, such as by dry heat. The dry heattreatment is usually performed at temperatures between 100° C. and 250°C., such as about 110° C. to about 200° C. In particular the temperaturemay be in the range of 110-160° C., e.g. in the range of 110-140° C., orin the range of 120-180° C., or in the range of 130-170° C., or in therange of 130-160° C., or in the range of 120-150° C. The period of timefor cross-linking may be optimised by a skilled person and is normally aperiod between about 10 minutes to about 12 hours, such as about 1 hourto about 10 hours, for example between about 2 hours to about 10 hours,such as between about 4 hours to about 8 hours, for example betweenabout 5 hours to about 7 hours, such as about 6 hours.

Examples of suitable chemical cross-linking agents include but are notlimited to aldehydes, in particular glutaraldehyde and formaldehyde,acyl azide, caboiimides, hexamethylene diisocyanate, polyether oxide,1,4-butanedioldiglycidyl ether, tannic acid, aldose sugars, e.g.D-fructose, genipin and dye-mediated photo-oxidation. Specific compoundsinclude but are not limitedtol-(3-dimethylaminopropyl)-3-ethylcarboiimide hydrochloride (EDC),dithiobis(propanoic dihydrazide) (DTP),I-ethyl-3-(3-dimethylamino-propyl)-carbodiimide (EDAC).

In one embodiment the biocompatible polymer in powder form is obtainedfrom a cross-linked sponge. The biocompatible polymer may for example beobtained from cross-linked sponges of gelatine or collagen, inparticular cross-linked sponges of gelatine (such as the commerciallyavailable Spongostan® sponges and Surgifoam® sponges). The cross-linkedsponges are micronised by methods known in the art to obtain across-linked biocompatible polymer in powder form, such as by rotarybed, extrusion, granulation and treatment in an intensive mixer, ormilling (e.g. by using a hammer mill or a centrifugal mill).

Spongostan®/Surgifoam® available from Ethicon is a gelatine basedcross-linked absorbable haemostatic sponge. It absorbs >35 g of blood/gand within 4-6 weeks it is completely absorbed in the human body.

The cross-linked powder particles are in one embodiment less thanapproximately 1000 microns in size, i.e. so that they are able to passthrough a 1×1 mm sieve.

In one embodiment, the paste prior to drying comprises between about 10%to about 60% of the biocompatible polymer, for example about 10% toabout 50% of the biocompatible polymer, such as about 10% to about 40%of the biocompatible polymer, for example about 10% to about 30% of thebiocompatible polymer, such as about 12% to about 25% of thebiocompatible polymer, for example about 14% to about 25% of thebiocompatible polymer, such as about 15% to about 25% of thebiocompatible polymer, for example about 16% to about 20% of thebiocompatible polymer, such as about 17% to about 20% of thebiocompatible polymer, for example about 18% to about 20% of thebiocompatible polymer.

In one embodiment, the paste prior to drying comprises more than 10% ofthe biocompatible polymer, such as more than 15% of the biocompatiblepolymer, for example more than 16% of the biocompatible polymer, such asmore than 17% of the biocompatible polymer, for example more than 18% ofthe biocompatible polymer, such as more than 19% of the biocompatiblepolymer, for example more than 20% of the biocompatible polymer.

In one embodiment, the paste prior to drying comprises less than 40% ofthe biocompatible polymer, such as less than 30% of the biocompatiblepolymer, for example less than 25% of the biocompatible polymer, such asless than 20% of the biocompatible polymer.

In a preferred embodiment, the paste prior to drying comprises betweenabout 10% to about 30% of the biocompatible polymer, more preferredbetween about 15% to about 25% of the biocompatible polymer, such asabout 20% of the biocompatible polymer.

In one embodiment, the paste prior to drying comprises between about 15%to about 20% of the biocompatible polymer, such as between about 16% toabout 20% of the biocompatible polymer, for example between about 17%and about 20% of the biocompatible polymer, such as between about 18% toabout 20% of the biocompatible polymer.

In one embodiment, the paste prior to drying comprises between about 20%to about 25% of the biocompatible polymer, such as between about 21% toabout 25% of the biocompatible polymer, for example between about 22%and about 25% of the biocompatible polymer, such as between about 23% toabout 25% of the biocompatible polymer.

After drying, the composition comprises between about 40% and 80% of thebiocompatible polymer, such as between about 45% and 80% of thebiocompatible polymer, for example between about 50% and 80% of thebiocompatible polymer, such as between about 55% and 80% of thebiocompatible polymer.

In one embodiment, the composition after drying comprises between about40% and 80% of the biocompatible polymer, such as between about 45% and75% of the biocompatible polymer, for example between about 50% and 70%of the biocompatible polymer.

In one embodiment, the dry composition of the present disclosurecomprises more than about 30% of the biocompatible polymer, such as morethan about 40% of the biocompatible polymer, for example more than about45% of the biocompatible polymer, such as more than about 50% of thebiocompatible polymer, for example more than about 55% of thebiocompatible polymer, such as more than about 60% of the biocompatiblepolymer, for example more than about 65% of the biocompatible polymer,such as more than about 70% of the biocompatible polymer, for examplemore than about 75% of the biocompatible polymer, such as more thanabout 80% of the biocompatible polymer.

In one embodiment, the dry composition of the present disclosurecomprises less than about 80% of the biocompatible polymer, such as lessthan about 70% of the biocompatible polymer, for example less than about65% of the biocompatible polymer, such as less than about 60% of thebiocompatible polymer, for example less than about 55% of thebiocompatible polymer, such as less than about 50% of the biocompatiblepolymer.

Aqueous Medium

The aqueous medium of the present disclosure may be any aqueous mediumsuitable for preparing a paste suitable for haemostatic use known to aperson of skill, e.g. water, saline, a calcium chloride solution or abuffered aqueous medium. The water may be WFI (Water For Injection). Theaqueous medium is selected so that the tonicity of the reconstitutedpaste product is suitable for use on a human or animal subject.

The aqueous medium of the present disclosure is in one embodiment asaline solution.

The aqueous medium of the present disclosure is in one embodiment acalcium chloride solution.

In other embodiments, the aqueous medium is water.

The aqueous medium may also be a buffered aqueous medium suitable foruse in a haemostatic paste. Any suitable buffering agent known to aperson of skill may be used, such as one or more buffering agentsselected from the group consisting of:

Sodium citrate; Citric acid, Sodium citrate; Acetic acid, Sodiumacetate; K2HPO4, KH2PO4; Na2HPO4, NaH2PO4; CHES; Borax, Sodiumhydroxide; TAPS; Bicine; Tris; Tricine; TAPSO; HEPES; TES; MOPS; PIPES;Cacodylate; SSC; MES, or others. The pH of the buffered aqueous mediumshould be suitable for creating a haemostatic paste intended for humanuse and can be determined by the skilled person.

The amount of aqueous medium is adjusted to the amount of biocompatiblepolymer so that a paste of a suitable consistency forms. To optimisemanufacturing, it may in some instances be desirable to add less waterto the paste prior to drying than is desired in the final reconstitutedpaste composition so that the paste takes up as little space as possibleand less energy is required to remove water in the drying step. Thepaste prior to drying may thus have lower water content than the finalreconstituted paste. The “missing” water can be added duringreconstitution to obtain a flowable paste having a desiredwater:biocompatible polymer ratio.

The paste of the present disclosure prior to drying comprises betweenabout 50% and about 90% of water, such as between about 55% and about85% of water, for example between about 60% and about 80% of water, suchas about 70% of water.

Preferably, the paste of the present disclosure prior to dryingcomprises between about 60% and about 80%, more preferred about 70% toabout 75% of water.

After drying, the dry composition comprises less than about 5% of water,such as less than about 3% of water, preferably less than about 2% ofwater, more preferred less than about 1.5% of water, even more preferredless than about 1% of water or even less. Hence, in one embodiment, thedry composition comprises from about 0.1 to about 5% water, such as fromabout 0.1% to about 2% water.

In one embodiment, the residual water content in the dry composition isabout 0.5% or less. Such a low residual water content is possible withe.g. industrial freeze-drying apparatuses.

A low residual water content in the composition after drying isdesirable as it decreases the risk of microbial growth in the drycomposition. Furthermore, a low residual water content is essential ifthe composition comprises bioactive agents that are unstable in aqueousconditions, such as e.g. thrombin. If thrombin is present in thecomposition of the present disclosure, the residual water content in thedry composition is preferably less than about 3% water, more preferredless than about 2% water.

Polyols

According to the present disclosure, one or more polyols are added tothe composition prior to drying the composition. The one or more polyolsplay a role in achieving a dry composition which upon addition of aliquid in the form of an aqueous medium such as water spontaneouslyreconstitutes to form a paste of an optimal consistency for haemostaticpurposes without the use of mechanical mixing or stirring of any kind.

A polyol as defined herein is a compound with multiple hydroxylfunctional groups. Polyols as defined herein include sugars (mono-, di-and polysaccharides) and sugar alcohols and derivatives thereof.

Monosaccharides include but are not limited to glucose, fructose,galactose, xylose and ribose.

Disaccharides include but are not limited to sucrose (saccharose),lactulose, lactose, maltose, trehalose and cellobiose.

Polysaccharides include but are not limited to starch, glycogen,cellulose and chitin.

A sugar alcohol, also known as a polyalcohol is a hydrogenated form ofcarbohydrate, whose carbonyl group (aldehyde or ketone, reducing sugar)has been reduced to a primary or secondary hydroxyl group (hence thealcohol). Sugar alcohols have the general formula H(HCHO)_(n+1)H,whereas sugars have H(HCHO)_(n)HCO. Some common sugar alcohols which maybe used in the method of the present disclosure include but are notlimited to: Glycol (2-carbon), Glycerol (3-carbon), Erythritol(4-carbon), Threitol (4-carbon), Arabitol (5-carbon), Xylitol(5-carbon), Ribitol (5-carbon), Mannitol (6-carbon), Sorbitol(6-carbon), Dulcitol (6-carbon), Fucitol (6-carbon), Iditol (6-carbon),Inositol (6-carbon; a cyclic sugar alcohol), volemitol (7-carbon),Isomalt (12-carbon), Maltitol (12-carbon), Lactitol (12-carbon),Polyglycitol.

In one embodiment, the composition comprises a single polyol.

In one embodiment of the disclosure, the composition comprises more thanone polyol, such as two, three, four, five, six or even more differentpolyols.

In one embodiment of the disclosure, the composition comprises twopolyols, for example mannitol and glycerol or trehalose and a glycol.

In one embodiment of the disclosure, the composition comprises one ormore sugar alcohols, such as one or more sugar alcohols selected fromthe group consisting of Glycol, Glycerol, Erythritol, Threitol,Arabitol, Xylitol, Ribitol, Mannitol, Sorbitol, Dulcitol, Fucitol,Iditol, Inositol, volemitol, Isomalt, Maltitol, Lactitol andPolyglycitol.

In one embodiment, the composition comprises one or more sugar alcoholsand one or more sugars, such as one sugar alcohol and one sugar.

In one embodiment, the composition comprises one sugar alcohol andoptionally one or more additional polyols, which may be either sugaralcohols or sugars.

In one embodiment, the composition does not comprise a sugar as the onlypolyol.

In one embodiment of the disclosure, the composition comprises mannitol.

In one embodiment of the disclosure, the composition comprises sorbitol.

In one embodiment of the disclosure, the composition comprises glycerol.

In one embodiment of the disclosure, the composition comprisestrehalose.

In one embodiment of the disclosure, the composition comprises glycol,such as propylene glycol.

In one embodiment of the disclosure, the composition comprises xylitol.

In one embodiment of the disclosure, the composition comprises maltitol.

In one embodiment of the disclosure, the composition comprises sorbitol.

In one embodiment the paste prior to drying comprises from about 1% toabout 40% of one or more polyols, for example from about 1% to about 30%of one or more polyols, such as from about 1% to about 25% of one ormore polyols, for example from about 1% to about 20% of one or morepolyols, such as from about 1% to about 15% of one or more polyols, suchas from about 1% to about 14% of one or more polyols, for example fromabout 1% to about 13% of one or more polyols, such as from about 1% toabout 12% of one or more polyols, for example from about 1% to about 11%of one or more polyols, such as about 1% to about 10% of one or morepolyols.

In one embodiment the paste prior to drying comprises from about 2% toabout 40% of one or more polyols, for example from about 2% to about 30%of one or more polyols, such as from about 2% to about 25% of one ormore polyols, for example from about 2% to about 20% of one or morepolyols, such as from about 2% to about 18% of one or more polyols, forexample from about 2% to about 17% of one or more polyols, such as fromabout 2% to about 16% of one or more polyols, for example from about 2%to about 15% of one or more polyols, such as from about 2% to about 14%of one or more polyols, for example from about 2% to about 13% of one ormore polyols, such as from about 2% to about 12% of one or more polyols,for example from about 2% to about 11% of one or more polyols, such asabout 2% to about 10% of one or more polyols.

In one embodiment the paste prior to drying comprises from about 3% toabout 40% of one or more polyols, for example from about 3% to about 30%of one or more polyols, such as from about 3% to about 25% of one ormore polyols, for example from about 3% to about 20% of one or morepolyols, such as from about 3% to about 18% of one or more polyols, forexample from about 3% to about 17% of one or more polyols, such as fromabout 3% to about 16% of one or more polyols, for example from about 3%to about 15% of one or more polyols, such as from about 3% to about 14%of one or more polyols, for example from about 3% to about 13% of one ormore polyols, such as from about 3% to about 12% of one or more polyols,for example from about 3% to about 11% of one or more polyols, such asabout 3% to about 10% of one or more polyols.

In one embodiment, the paste prior to drying comprises more than about5% of one or more polyols, hence in one embodiment the paste prior todrying comprises from about 5% to about 40% of one or more polyols, forexample from about 5% to about 30% of one or more polyols, such as fromabout 5% to about 25% of one or more polyols, for example from about 5%to about 20% of one or more polyols, such as from about 5% to about 18%of one or more polyols, for example from about 5% to about 17% of one ormore polyols, such as from about 5% to about 16% of one or more polyols,for example from about 5% to about 15% of one or more polyols, such asfrom about 5% to about 14% of one or more polyols, for example fromabout 5% to about 13% of one or more polyols, such as from about 5% toabout 12% of one or more polyols, for example from about 5% to about 11%of one or more polyols, such as about 5% to about 10% of one or morepolyols.

In one embodiment the paste prior to drying comprises from about 6% toabout 40% of one or more polyols, for example from about 6% to about 30%of one or more polyols, such as from about 6% to about 25% of one ormore polyols, for example from about 6% to about 20% of one or morepolyols, such as from about 6% to about 18% of one or more polyols, forexample from about 6% to about 17% of one or more polyols, such as fromabout 6% to about 16% of one or more polyols, for example from about 6%to about 15% of one or more polyols, such as from about 6% to about 14%of one or more polyols, for example from about 6% to about 13% of one ormore polyols, such as from about 6% to about 12% of one or more polyols,for example from about 6% to about 11% of one or more polyols, such asabout 6% to about 10% of one or more polyols.

In one embodiment the paste prior to drying comprises from about 10% toabout 40% of one or more polyols, for example from about 10% to about30% of one or more polyols, such as from about 10% to about 25% of oneor more polyols, for example from about 10% to about 20% of one or morepolyols, such as from about 10% to about 18% of one or more polyols, forexample from about 10% to about 17% of one or more polyols, such as fromabout 10% to about 16% of one or more polyols, for example from about10% to about 15% of one or more polyols.

In one embodiment, the paste prior to drying comprises more than about1% of one or more polyols, such as more than about 2% of one or morepolyols, for example more than about 3% of one or more polyols, such asmore than about 4% of one or more polyols, for example more than about5% of one or more polyols, such as more than about 6% of one or morepolyols, for example more than about 7% of one or more polyols, such asmore than about 8% of one or more polyols, for example more than about9% of one or more polyols, such as more than about 10% of one or morepolyols.

In one embodiment, the paste prior to drying comprises less than about20% of one or more polyols, such as less than about 18% of one or morepolyols, for example less than about 17% of one or more polyols, such asless than about 16% of one or more polyols, for example less than about15% of one or more polyols, such as less than about 14% of one or morepolyols, for example less than about 13% of one or more polyols, such asless than about 12% of one or more polyols, for example less than about11% of one or more polyols, such as less than about 10% of one or morepolyols.

After drying, the dry composition comprises from about 10% to about 60%of one or more polyols, such as from about 20% to about 50% of one ormore polyols, such as from about 20% to about 45% of one or morepolyols, for example from about 20% to about 40%, such as from about 20%to about 35% of one or more polyols, for example from about 20% to about30% of one or more polyols.

In one embodiment, the dry composition comprises from about 20% to about60% of one or more polyols, such as from about 20% to about 50% of oneor more polyols, for example from about 20% to about 50%, such as fromabout 20% to about 45% of one or more polyols, for example from about20% to about 40%, such as from about 20% to about 30% of one or morepolyols.

In one embodiment, the dry composition comprises from about 25% to about60% of one or more polyols, such as from about 25% to about 50% of oneor more polyols, for example from about 25% to about 45% of one or morepolyols, such as from about 25% to about 40% of one or more polyols, forexample from about 25% to about 35% of one or more polyols, such as fromabout 25% to about 30% of one or more polyols.

In one embodiment, the dry composition comprises from about 27% to about60% of one or more polyols, such as from about 27% to about 50% of oneor more polyols, for example from about 27% to about 45% of one or morepolyols, such as from about 27% to about 40% of one or more polyols, forexample from about 27% to about 35% of one or more polyols, such as fromabout 27% to about 30% of one or more polyols.

In one embodiment, the dry composition comprises from about 30% to about60% of one or more polyols, such as from about 30% to about 50% of oneor more polyols, for example from about 30% to about 45% of one or morepolyols, such as from about 30% to about 40% of one or more polyols, forexample from about 30% to about 35% of one or more polyols.

In one embodiment, the dry composition comprises less polyol thanbiocompatible polymer, i.e. the polyol:biocompatible polymer ratio isless than 1:1, such as less than or about 0.9:1, for example less thanor about 0.8:1, such as less than or about 0.7:1, for example less thanor about 0.6:1, such as less than or about 0.5:1, such as less than orabout 0.4:1, for example less than or about 0.3:1, such as less than orabout 0.2:1, for example less than or about 0.1:1. Thepolyol:biocompatible polymer ratio is the same in the paste prior todrying.

In one embodiment, the polyol:biocompatible polymer ratio is betweenabout 0.1:1 and 1:1; such as between about 0.2:1 and 1:1, for examplebetween about 0.3:1 and 1:1, such as between about 0.4:1 and 1:1.

In a preferred embodiment, the polyol:biocompatible polymer ratio isbetween about 0.2:1 and 0.8:1; such as between about 0.2:1 and 0.7:1,for example between about 0.2:1 and 0.6:1, such as between about 0.2:1and 0.5:1. Even more preferred, the polyol:biocompatible polymer ratiois between about 0.3:1 and 0.8:1; such as between about 0.3:1 and 0.7:1,for example between about 0.3:1 and 0.6:1, such as between about 0.3:1and 0.5:1, for example between about 0.35:1 and 0.5:1, such as betweenabout 0.35:1 and 0.45:1.

In one embodiment the polyol is not polyethylene glycol.

Extrusion Enhancer

It has previously been shown that the provision of certain extrusionenhancers, such as albumin in an appropriate amount, enables the use ofhigher gelatine concentrations as it decreases the amount of forceneeded to extrude the gelatine paste composition from e.g. a syringe.The use of higher gelatine concentrations may in turn improve thehaemostatic properties of such products. It is necessary to provide theextrusion enhancers in appropriate amounts. The amounts shall be highenough so as to obtain the extrusion effect, i.e. to enable a flowablepaste even for relatively high amounts of the biocompatible polymer,e.g. cross-linked gelatine, so that the haemostatic paste compositioncan be accurately applied by a surgeon using e.g. a syringe comprisingan applicator tip; on the other hand, the amounts shall be as low as toprevent negative functional properties of the haemostatic composition.

In a preferred embodiment, the extrusion enhancer is albumin, especiallyhuman serum albumin.

In the wet paste composition before drying, the extrusion enhancer, suchas albumin, is preferably present in an amount of between about 0.1% toabout 10%, such as between about 0.2% to about 8%, for example betweenabout 0.3% to about 7%, preferably between about 0.5% to about 5%, morepreferred between about 1% to about 4%.

In the dry composition, the extrusion enhancer such as albumin ispreferably present in an amount of between about 0.3% to about 30%, suchas between about 0.5% to about 25%, for example between about 1% toabout 20%, preferably between about 2% to about 15%.

In one embodiment, the extrusion enhancer is not present in the drycomposition, but is instead introduced into the paste composition duringreconstitution. For example the extrusion enhancer may be present in theaqueous medium used for reconstitution of the paste, thereby obtaining awet paste composition comprising the extrusion enhancer.

In one embodiment, the reconstituted wet paste composition being presentin an applicator device suitable for dispensing compositions in pasteform, such as a syringe, has a mean extrusion force of 40 N or below,preferably below 35 N, especially preferred below 30 N or even below 20N. Extrusion force may be tested by suitable methods known in the artcapable of measuring the force required to extrude a paste product froma syringe.

In one embodiment, the reconstituted wet paste composition has a meanextrusion force (employing the test method described in example 1 of WO2013/060770) of 40 N or below, preferably below 35 N, especiallypreferred below 30 N or even below 20 N.

Another class of extrusion enhancers according to the present disclosureare phospholipids, such as phosphatidylcholine and -serine, or complexmixtures such as lecithins or soy bean oils.

Bioactive Agent

In one embodiment of the present disclosure, the dry compositioncomprises one or more bioactive agents. It is essential that thebioactive agent retains its bioactivity, i.e. that the bioactive agentis biologically active in the paste after reconstitution of the drycomposition. Many bioactive agents are unstable in solution,particularly enzymes and other proteins that may be degraded or losetheir secondary structure when water is present.

In one embodiment the bioactive agent stimulates wound healing and/orhaemostasis, such as thrombin.

Conventionally, a thrombin solution is added to a gelatine powder tomake a haemostatic paste directly at the surgical site at the time ofneed of the haemostatic paste, e.g. by using commercially availablehaemostatic kits such as Floseal and Surgiflo®. The thrombin solutionmust be made just prior to making the paste as thrombin in solution isvery unstable and will self-degrade rapidly. The making of a thrombinsolution at the surgical site is time consuming and involves a risk ofmaking mistakes regarding the correct dilution of thrombin.

The method of the present disclosure allows for the addition of thrombinto a paste prior to drying, thereby resulting in a dry compositioncomprising thrombin, which upon reconstitution with a suitable aqueousmedium, such as water, will comprise a desired amount of thrombinwithout the need for time-consuming and error-prone thrombin dilutionsteps and addition at the surgical site. That thrombin may be includedin the dry composition constitutes a clear advantage over conventionalmethods for making haemostatic pastes.

The present inventor has previously shown that thrombin may be includedin a paste and dried by freeze-drying according with essentially no lossof thrombin activity measured in the reconstituted paste.

Thrombin may be added to the paste prior to drying at a concentration inthe range of about 100 IU/ml paste to about 500 IU/ml paste, such asabout 150 IU/ml paste to about 450 IU/ml paste, for example about 200IU/ml paste to about 400 IU/ml paste, such as about 250 IU/ml paste toabout 350 IU/ml paste.

In one embodiment, the one or more bioactive agents can be e.g. thrombinor thrombin in combination with fibrinogen, or thrombin and fibrinogenin combination with Factor XIII, or thrombin and fibrinogen and FactorXIII in combination with tranexamic acid.

Thrombin is a “trypsin-like” serine protease protein that in humans isencoded by the F2 gene. Prothrombin (coagulation factor II) isproteolytically cleaved to form thrombin in the coagulation cascade,which ultimately results in the stemming of blood loss. Thrombin in turnacts as a serine protease that converts soluble fibrinogen intoinsoluble strands of fibrin, as well as catalyzing many othercoagulation-related reactions. In the blood coagulation pathway,thrombin acts to convert factor XI to Xla, VIII to Villa, V to Va, andfibrinogen to fibrin.

A preferred bioactive agent is thrombin. In one embodiment, the thrombinis added as prothrombin.

In one embodiment, the dry composition comprises one or more bioactiveagents that stimulate bone and/or tendon healing such as one or moregrowth factors selected from the group consisting of matrixmetalloproteinases (MMPs), insulin-like growth factor 1 (IGF-I),platelet-derived growth factor (PDGF), vascular endothelial growthfactor (VEGF), basic fibroblast growth factor (bFGF) and transforminggrowth factor beta (TGF-β).

In one embodiment, the dry composition comprises one or more BoneMorphogenetic Proteins (BMPs). Bone morphogenetic proteins (BMPs) are asubgroup of the TGF-β superfamily. Bone Morphogenetic Proteins (BMPs)are a group of growth factors also known as cytokines and asmetabologens. Originally discovered by their ability to induce theformation of bone and cartilage, BMPs are now considered to constitute agroup of pivotal morphogenetic signals, orchestrating tissuearchitecture throughout the body.

In one embodiment, the dry composition comprises one or more matrixmetalloproteinases (MMPs). MMPs are zinc-dependent endopeptidases. MMPshave a very important role in the degradation and remodeling of theextracellular matrix (ECM) during the healing process after an injury.Certain MMPs including MMP-1, MMP-2, MMP-8, MMP-13, and MMP-14 havecollagenase activity, meaning that, unlike many other enzymes, they arecapable of degrading collagen I fibrils.

These growth factors all have different roles during the healingprocess. IGF-1 increases collagen and proteoglycan production during thefirst stage of inflammation, and PDGF is also present during the earlystages after injury and promotes the synthesis of other growth factorsalong with the synthesis of DNA and the proliferation of cells. Thethree isoforms of TGF-β (TGF-β1, TGF-β2, TGF-β3) are known to play arole in wound healing and scar formation. VEGF is well known to promoteangiogenesis and to induce endothelial cell proliferation and migration.

In one embodiment, the dry composition comprises flakes or particles ofextracelluar matrix (ECM). ECM is the extracellular part of animaltissue that usually provides structural support to the animal cells inaddition to performing various other important functions. ECM has beenshown to have very beneficial effect in healing as it facilitatesfunctional tissue regeneration.

The variety of biological agents that can be used in conjunction withthe paste of the disclosure is vast. In general, biological agents whichmay be administered via the compositions disclosed herein include,without limitation, antiinfectives, such as antibiotics and antiviralagents; analgesics and analgesic combinations; antihelmintics;antiarthritics; anticonvulsants; antidepressants; antihistamines;antiinflammatory agents; antimigraine preparations; antineoplastics;antiparkinsonism drugs; antipsychotics; antipyretics, antispasmodics;anticholinergics; sympathomimetics; xanthine derivatives; cardiovascularpreparations including calcium channel blockers and beta-blockers suchas pindolol and antiarrhythmics; antihypertensives; diuretics;vasodilators, including general coronary, peripheral and cerebral;central nervous system stimulants; hormones, such as estradiol and othersteroids, including corticosteroids; immunosuppressives; musclerelaxants; parasympatholytics; psychostimulants; naturally derived orgenetically engineered proteins, polysaccharides, glycoproteins, orlipoproteins; oligonucleotides, antibodies, antigens, cholinergics,chemotherapeutics, radioactive agents, osteoinductive agents,cystostatics heparin neutralizers, procoagulants and hemostatic agents,such as prothrombin, thrombin, fibrinogen, fibrin, fibronectin,heparinase, Factor X/Xa, Factor VII/VIIa, Factor VIII/VIIIa, FactorIX/IXa, Factor XI/Xla, Factor XII/XlIa, Factor XIII/XIIIa, tissuefactor, batroxobin, ancrod, ecarin, von Willebrand Factor, collagen,elastin, albumin, gelatin, platelet surface glycoproteins, vasopressin,vasopressin analogs, epinephrine, selectin, procoagulant venom,plasminogen activator inhibitor, platelet activating agents andsynthetic peptides having haemostatic activity.

Further Compounds

The dry composition of the present disclosure may further comprise oneor more of the following: DMSO (dimethyl sulfoxide),2-Methyl-2,4-pentanediol (MPD) and/or one or more of the compoundsmentioned in the table below.

Bulking agent Buffering agent Solubilising agent MiscellaneousSugars/Sugar Citric acid Complexing Tonicifying alcohols: Sodium citrateagent: agent: Mannitol Potassium citrate Ethylediamine Sodium LactoseTartaric acid tetra acetic acid chloride Sucrose Sodium (EDTA) SucroseTrehalose phosphate Alpha cyclo- Mannitol Sorbitol Tris base dextrinDextrose Glucose Tris HCl Hydroxypropyl-β- Raffinose Tris acetatecyclodextrin Zinc chloride (HP-β-CD) Sodium acetate Potassium acetateArginine Amino acids: pH adjusting agent: Surfactants: AntimicrobialArginine Hydrochloric acid polysorbate 80 agents: Glycine Sodiumhydroxide Benzalkonium Histidine Meglumine chloride benzyl alcoholphenol m-cresol methyl paraben ethyl paraben Polymer: Co-solvents:Collapse Dextran Tert-butyl alcohol temperature Polyethylene Iso-propylalcohol modifier: glycol Dichloromethane Dextran Ethanol HydroxyethylAcetone starch Glycerol Ficoll gelatin

In one embodiment, the dry composition of the present disclosurecomprises one or more antimicrobial agents, such as one or moreantibacterial agents.

In one embodiment, the dry composition of the present disclosurecomprises benzalkonium chloride (BAC).

In one embodiment, the dry composition of the present disclosure doesnot comprise an antimicrobial agent.

Making a Paste

According to the method of the present disclosure, the biocompatiblepolymer, the extrusion enhancer and the one or more polyols are mixedwith a suitable aqueous medium to obtain a paste. The mixing may beperformed in any suitable way known to a person of skill, e.g. by mixingthe contents manually or by using an electrical mixing apparatus, suchas a hand mixer, a kitchen mixer or an industrial mixer.

Mixing of the paste can generally be performed at room temperature(20-25° C.). However, if thrombin or other enzymes are included in thepaste, it is advisable to perform the mixing of the paste at chilledtemperatures and/or within a short time period to avoid or decrease theproteolytic activity of thrombin, as it is well-known that thrombin isliable to self-degradation in solution. Hence, when thrombin or otherproteolytic enzymes are to be included in the paste, the mixing of thepaste may be performed at temperatures below room temperature, such asat about 2° C. to about 20° C., for example at about 2° C. to about 15°C., preferably at about 4° C.

Another or an additional way of preserving the thrombin bioactivity inthe paste is to keep the time that thrombin is in a wet state, i.e. themixing time, at a minimum. Hence, when thrombin or other proteolyticenzymes are to be included in the paste, the mixing of the paste isusually performed within about 5 minutes to about 10 hours, such asabout 5 minutes to about 5 hours, for example about 5 minutes to about 2hours, preferably about 5 minutes to about 1 hour.

The present inventor has previously found that it is not essential toperform the mixing of the paste at low temperatures to avoid loss ofthrombin activity as no decrease in thrombin activity was discoveredwhen mixing of the paste was performed at ambient temperatures.

Containers

Any suitable container known to a person of skill may be used forpreparing the paste and holding the paste while drying, such as vials,jars, tubes, trays, cartridges or syringes.

In one embodiment, the paste is prepared in one container andtransferred to another container for drying, wherein said othercontainer may be selected from a vial, a jar, a tube, a tray, acartridge and a syringe.

A “jar” according to the present disclosure is a rigid, approximatelycylindrical container with a wide mouth opening. Jars may comprise are-closable closure unit/lid applied to the mouth of the jar.

The containers may be made from any suitable material such as glass,ceramic, plastic or metal, such as stainless steel.

Examples of suitable plastic materials include but are not limited topolyethylene, polypropylene, polystyrene, polyvinyl chloride, andpolytetrafluoroethylene (PTFE).

In one embodiment, the paste is filled into and dried within a syringeor other known applicators suitable for dispensing flowable haemostaticcompositions.

In one embodiment, the paste is dried within an applicator suitable fordispensing compositions in paste form, such as a syringe. Thus, in oneembodiment, the present disclosure relates to an applicator, such as asyringe, comprising a dry composition contained therein.

The dry composition of the present disclosure may be prepared in variousshapes, forms and sizes depending on the shape of the container used.They may be e.g. in the form of plugs, disks, rods, tubes, conicalcylinders, sheets, spheres, half spheres, tablets, pellets, granules, oreven fine particulates or powders (pulverised).

Haemostatic Sheet

In one embodiment the dry composition is in the form of a sheet, i.e. asubstantially flat composition.

A dry composition in the form of a sheet may be obtained by spreadingthe paste thinly and evenly on a surface followed by drying of the pasteto obtain a substantially flat dry sheet composition. A dry compositionin the form of a sheet will upon contact with a liquid reconstitutespontaneously to form a paste. Thus, a dry composition in the form of asheet has the advantages of both traditionally used surgical sponges inthat it can cover relatively large areas and the advantage of a paste inthat it, when wetted, conforms easily to uneven surfaces upon wetting.

The dry composition in the form of a sheet is soft and flexible.

In one embodiment the present disclosure relates to a dry composition inthe form of a sheet for use in haemostasis and/or wound healing.

In one embodiment, the sheet is not pre-wetted before use, i.e. beforeapplication to a wound.

The height of the dry sheet composition is in one embodiment betweenabout 0.5 mm and about 10 mm, preferably between about 1 mm and 5 mm,more preferred between about 1 mm and 3 mm, such as about 2 mm.

The size (width and depth) of the dry sheet composition depends on theintended use of the sheet and can be selected by the skilled person. Thedry sheet material may e.g. be rectangular, square or circular. Forexample, the dry sheet composition may e.g. be in the form of arectangle of approximately 5 cm×10 cm, 2 cm×6 cm, 6 cm×8 cm or 8 cm×12cm.

In one embodiment, the dry sheet composition is cut into the desiredshape prior to use.

Drying the Paste

According to the present disclosure the paste is dried to obtain the drycomposition. The paste may be dried by any suitable methods known to aperson of skill. Examples of suitable drying methods includefreeze-drying and spray drying.

In one embodiment, the paste is frozen prior to the drying step.

In a preferred embodiment, the paste is freeze-dried. Any suitablefreeze-drying technique and equipment known to the person of skill maybe used.

Freeze-drying (also known as lyophilisation and cryodesiccation) is adehydration process typically used to preserve a perishable material ormake the material more convenient for transport. Freeze-drying works byfreezing the material and then reducing the surrounding pressure toallow the frozen water in the material to sublimate directly from thesolid phase to the gas phase.

There are essentially three categories of freeze-dryers: the manifoldfreeze-dryer, the rotary freeze-dryer and the tray style freeze-dryer.Two components are common to all types of freeze-dryers: a vacuum pumpto reduce the ambient gas pressure in a vessel containing the substanceto be dried and a condenser to remove the moisture by condensation on asurface cooled to −40 to −80° C. The manifold, rotary and tray typefreeze-dryers differ in the method by which the dried substance isinterfaced with a condenser. In manifold freeze-dryers a short usuallycircular tube is used to connect multiple containers with the driedproduct to a condenser. The rotary and tray freeze-dryers have a singlelarge reservoir for the dried substance.

Rotary freeze-dryers are usually used for drying pellets, cubes andother pourable substances. The rotary dryers have a cylindricalreservoir that is rotated during drying to achieve a more uniform dryingthroughout the substance. Tray style freeze-dryers usually haverectangular reservoir with shelves on which products, such aspharmaceutical solutions and tissue extracts, can be placed in trays,vials and other containers.

Manifold freeze-dryers are usually used in a laboratory setting whendrying liquid substances in small containers and when the product willbe used in a short period of time. A manifold dryer will dry the productto less than 5% moisture content. Without heat, only primary drying(removal of the unbound water) can be achieved. A heater must be addedfor secondary drying, which will remove the bound water and will producea lower moisture content.

Tray style freeze-dryers are typically larger than the manifold dryersand are more sophisticated. Tray style freeze-dryers are used to dry avariety of materials. A tray freeze-dryer is used to produce the driestproduct for long-term storage. A tray freeze-dryer allows the product tobe frozen in place and performs both primary (unbound water removal) andsecondary (bound water removal) freeze-drying, thus producing the driestpossible end-product. Tray freeze-dryers can dry products in bulk or invials or other containers. When drying in vials, the freeze-drier issupplied with a stoppering mechanism that allows a stopper to be pressedinto place, sealing the vial before it is exposed to the atmosphere.This is used for long-term storage, such as vaccines.

Improved freeze drying techniques are being developed to extend therange of products that can be freeze dried, to improve the quality ofthe product, and to produce the product faster with less labour.

Ever since the 1930s, industrial freeze drying has been dependent on asingle type of equipment: the tray freeze drier. In 2005 a quicker andless-labour intensive freeze drying method was developed for bulkmaterials. This freeze drying process proved to be able to producefree-flowing powder from a single vessel. Known as [Active FreezeDrying] AFD technology, the new process used continuous motion toimprove mass transfer and hence cutting processing time, while alsoeliminating the need to transfer to and from drying trays and downstreamsize reduction devices.

There are four stages in the complete freeze-drying process:pre-treatment, freezing, primary drying, and secondary drying.

Pre-treatment includes any method of treating the product prior tofreezing. This may include concentrating the product, formulationrevision (i.e., addition of components to increase stability and/orimprove processing), decreasing a high vapor pressure solvent orincreasing the surface area. In many instances the decision to pre-treata product is based on theoretical knowledge of freeze-drying and itsrequirements, or is demanded by cycle time or product qualityconsiderations. Methods of pre-treatment include: Freeze concentration,Solution phase concentration, Formulation to Preserve.

Product Appearance, Formulation to Stabilize Reactive Products,Formulation to Increase the Surface Area, and Decreasing High VaporPressure Solvents.

In a lab, freezing is often done by placing the material in afreeze-drying flask and rotating the flask in a bath, called a shellfreezer, which is cooled by mechanical refrigeration, dry ice andmethanol, or liquid nitrogen. On a larger scale, freezing is usuallydone using a freeze-drying machine. In this step, it is important tocool the material below its triple point, the lowest temperature atwhich the solid and liquid phases of the material can co-exist. Thisensures that sublimation rather than melting will occur in the followingsteps. Larger crystals are easier to freeze-dry. To produce largercrystals, the product should be frozen slowly or can be cycled up anddown in temperature. This cycling process is called annealing. In othercases it is better that the freezing is done rapidly, in order to lowerthe material to below its eutectic point quickly, thus avoiding theformation of ice crystals. Usually, the freezing temperatures arebetween −40° C. and −80° C. The freezing phase is the most critical inthe whole freeze-drying process, because the product can be spoiled ifbadly done.

Amorphous materials do not have a eutectic point, but they do have acritical point, below which the product must be maintained to preventmelt-back or collapse during primary and secondary drying.

During the primary drying phase, the pressure is lowered (to the rangeof a few millibars or less), and enough heat is supplied to the materialfor the water to sublime. The amount of heat necessary can be calculatedusing the sublimating molecules' latent heat of sublimation. In thisinitial drying phase, about 95% of the water in the material issublimated. This phase may be slow (can be several days in theindustry), because, if too much heat is added, the material's structurecould be altered.

In this phase, pressure is controlled through the application of partialvacuum. The vacuum speeds sublimation, making it useful as a deliberatedrying process.

Furthermore, a cold condenser chamber and/or condenser plates provide asurface(s) for the water vapour to re-solidify on. This condenser playsno role in keeping the material frozen; rather, it prevents water vaporfrom reaching the vacuum pump, which could degrade the pump'sperformance. Condenser temperatures are typically below −50° C.

It is important to note that, in this range of pressure, the heat isbrought mainly by conduction or radiation; the convection effect isnegligible, due to the low air density.

The vapour pressure of water is the pressure at which water vapour issaturated. At higher pressures water would condense. The water vapourpressure is the partial pressure of water vapour in any gas mixturesaturated with water. The water vapour pressure determines thetemperature and pressure necessary for freeze-drying to occur.

Vapour pressure of water (mTorr=millitorr; mB=millibar)

Temp (C.) mTorr mB 0 4579 6.104 −4 3280 4.372 −8 2326 3.097 −12 16322.172 −16 1132 1.506 −20 930 1.032 −24 526 0.6985 −28 351 0.4669 −32 2310.3079 −36 150 0.2020 −40 96.6 0.1238 −44 60.9 0.0809 −48 37.8 0.0502−52 23.0 0.0300 −56 13.8 0.0183 −60 8.0 0.0107 −64 4.6 0.0061 −68 2.60.0034 −72 1.4 0.0018

The secondary drying phase aims to remove unfrozen water molecules,since the ice was removed in the primary drying phase. This part of thefreeze-drying process is governed by the material's adsorptionisotherms. In this phase, the temperature is raised higher than in theprimary drying phase, and can even be above 0° C., to break anyphysico-chemical interactions that have formed between the watermolecules and the frozen material. Usually the pressure is also loweredin this stage to encourage desorption (typically in the range ofmicrobars). However, there are products that benefit from increasedpressure as well.

After the freeze-drying process is complete, the vacuum may be brokenwith an inert gas, such as nitrogen, before the material is sealed.

At the end of the operation, the final residual water content in thefreeze-dried product is in general very low, such as around 2% or lower.

The freeze-drying process transforms the paste into a hard “cake-like”composition, which upon addition of an adequate amount of an aqueousmedium, such as water, will form a ready-to use paste spontaneously,i.e. no mechanical mixing/reconstitution is required for said paste toform.

In one embodiment, the hard cake-like structure obtained byfreeze-drying the paste is pulverised before addition of the aqueousmedium.

In an alternative embodiment, the dry composition of the presentdisclosure is obtained by spray-drying. Any spray drying technique andequipment known to the skilled person may be applied.

Spray drying is a method of producing a dry powder from a liquid orslurry by rapidly drying with a hot gas. Air is usually the heateddrying media; however, if the liquid is a flammable solvent such asethanol or the product is oxygen-sensitive then nitrogen is used.

All spray dryers use some type of atomizer or spray nozzle to dispersethe liquid or slurry into a controlled drop size spray. The most commonof these are rotary disks and single-fluid high pressure swirl nozzles.Alternatively, for some applications two-fluid or ultrasonic nozzles areused. Depending on the process needs, drop sizes from 10 to 500 μm canbe achieved with the appropriate choices. The most common applicationsare in the 100 to 200 μm diameter range. The dry powder obtained isoften free-flowing.

Spray dryers can dry a product very quickly compared to other methods ofdrying. They also turn a solution or slurry into a dried powder in asingle step, which can be advantageous for profit maximization andprocess simplification.

Outer Packaging

In one embodiment the dry composition contained within e.g. a syringe orother containment unit, is further contained within an outer packagingso that the dry product is kept sterile until use. This will allow theuser to remove the outer packaging and transfer the dry composition intoa sterile field. Here a suitable amount of aqueous medium can be added,whereupon a ready-to-use paste forms spontaneously within secondswithout any need for mechanical agitation, stirring or mixing.

The outer packaging is usually made from a flexible, semi-rigid or rigidmaterial and typically consists of materials such as plastic, aluminiumfoil and/or plastic laminate, where the plastic may be selected from thegroup consisting of PET, PETG, PE, LLDPE, CPP, PA, PETP, METPET, Tyvekand optionally bonded with an adhesive, such as polyurethane, orco-extruded.

In one embodiment, the outer packaging is an aluminium foil outerpackaging.

The outer packaging preferably forms a complete barrier to moisture.

The outer packaging is preferably able to endure sterilisation treatmentsuch as by radiation.

Sterilisation

The dry composition of the present disclosure is preferably sterile. Anysuitable sterilisation technique known in the art may be utilised. Thesterilisation preferably occurs after the packaging step, i.e. when thedry composition is contained within an outer packaging. Thus, in apreferred embodiment sterilisation is terminal sterilisation.

Sterilisation refers to any process that effectively kills or eliminatestransmissible agents (such as fungi, bacteria, viruses, prions and sporeforms etc.). Sterilisation of the dry composition can be achievedthrough e.g. application of heat, chemicals, and irradiation. Heatsterilization include autoclaving (uses steam at high temperatures) anddry heat; radiation sterilisation include X-rays, gamma and beta rays,UV light and subatomic particles; chemical sterilisation include usingethylene oxide gas, ozone, chlorine bleach, glutaraldehyde,formaldehyde, ortho phthalaldehyde, hydrogen peroxide and peraceticacid.

In one embodiment, the dry composition is sterilised by irradiation,e.g. ionizing irradiation, so as to provide sterility to thecomposition. Such irradiation may include e-beam (beta irradiation) orgamma irradiation. The level of irradiation and conditions forsterilisation, including the time that the composition is irradiated,are those that provide sterile compositions. Sterilisation conditionsare similar to those currently utilized in the preparation ofhaemostatic loose powders currently available. Once having the benefitof this disclosure, one skilled in the art will be able to readilydetermine the level of irradiation necessary to provide sterilecompositions.

When thrombin and/or other sensitive bioactive agents are present in thedried product, sterilisation is usually performed as terminalsterilisation with about 25 kGy or less of beta or gamma irradiation.

In one embodiment, sterilisation is performed with ethylene oxide.

Sterilisation with dry heat may typically be carried out by heating thedry composition to a temperature between 100° C. and 250° C., such asabout 110° C. to about 200° C. In particular the temperature may be inthe range of 110-160° C., e.g. in the range of 110-140° C., or in therange of 120-180° C., or in the range of 130-170° C., or in the range of130-160° C., or in the range of 120-150° C.

In one embodiment, the dry composition is not sterilised afterpackaging. When the dry composition is manufactured by asepticproduction techniques, the product is already sterile when placed in theouter packaging and no further sterilisation is required. Thus, in oneembodiment the present disclosure relates to a composition produced byaseptic techniques.

Reconstitution of the Dry Composition

The dry composition of the present disclosure may be reconstituted byaddition of a suitable amount of an aqueous medium. The aqueous mediumused to reconstitute the paste may e.g. be selected from water, saline,a CaCl₂) solution or a buffered aqueous solution.

In one embodiment, the aqueous medium used to reconstitute the drycomposition is water. In one embodiment, the tonicity of the aqueousmedium is selected so that the tonicity of the reconstituted paste iscompatible with use on human subjects in surgical procedures.

In one embodiment, the aqueous medium used to reconstitute the drycomposition is saline.

In one embodiment, the aqueous medium used to reconstitute the drycomposition comprises an extrusion enhancer, such as albumin. In thiscase, the dry composition preferably does not contain an extrusionenhancer.

Upon addition of the aqueous medium, a ready-to-use flowable paste formsspontaneously, i.e. within few seconds. Importantly, no mixing isrequired for the paste to form.

In one embodiment, the reconstituted paste contains a lowerconcentration of biocompatible polymer than the paste prior to dryingdue to the addition of more water than was removed from the paste duringthe drying step.

In one embodiment, the present disclosure relates to a method forreconstituting a dry composition comprising the steps of:

-   -   a. providing a dry composition as described herein, optionally        comprising an extrusion enhancer, and    -   b. adding an aqueous medium to the dry composition.

The aqueous medium may optionally comprise an extrusion enhancer such asalbumin.

The extrusion enhancer may be present in the dry composition and/or inthe reconstitution liquid, i.e. the aqueous medium.

In some embodiments the reconstituted paste comprises a similarconcentration of polymer, e.g. gelatine, as conventionally used flowablepaste compositions, but, due to the presence of the extrusion enhancer,the reconstituted paste of the present disclosure will be comparablyeasier to extrude from the applicator, e.g. a syringe.

In one embodiment, the reconstituted paste comprises a higher amount ofbiocompatible polymer than conventionally used flowable pasteformulations. The increased amount of polymer may lead to an improvedhaemostatic effect, while still allowing for facile extrusion of thepaste from e.g. a syringe.

In one embodiment, the reconstituted paste of the present disclosurecomprises between about 10% to about 25% of the biocompatible polymer,for example between about 10% to about 20% of the biocompatible polymer,such as between about 12% to about 18% of the biocompatible polymer, forexample between about 14% to about 16% of the biocompatible polymer,such as about 15% of the biocompatible polymer.

In one embodiment, the reconstituted paste of the present disclosurecomprises less than about 15% of the biocompatible polymer, for examplebetween about 10% to about 15% of the biocompatible polymer, such asbetween about 11% to about 15% of the biocompatible polymer, for examplebetween about 12% to about 15% of the biocompatible polymer, such asbetween about 13% to about 15% of the biocompatible polymer, for examplebetween about 14% to about 15% of the biocompatible polymer.

In one embodiment, the reconstituted paste of the present disclosurecomprises more than about 15% of the biocompatible polymer, such asbetween about 15% to about 25% of the biocompatible polymer, for examplebetween about 15% to about 20% of the biocompatible polymer, such asbetween about 16% to about 20% of the biocompatible polymer, for examplebetween about 17% to about 20% of the biocompatible polymer, such asbetween about 18% to about 20% of the biocompatible polymer.

Medical Use

The present disclosure further relates to use of the dry composition orthe paste obtained from the dry composition for promoting haemostasisand/or wound healing.

The paste of the present disclosure may e.g. be used in an array ofsurgical procedures wherein bleeding control is desired. A pasteconforms to irregular surfaces to stop bleeding fast and it is thereforeuseful for providing rapid haemostasis on rough or uneven surfaces wherehaemostatic sponges are not efficient.

Haemostatic pastes are prepared directly at the surgical site at thetime of need by the medical practitioner, i.e. doctors or nurses. Thepaste is thus often prepared under extremely stressful conditions and itis therefore essential that the process for preparing the paste issimple and fast to ensure that the bleeding is arrested as quickly aspossible and that no mistakes are made while preparing the paste. It isalso important that the consistency of the paste is suitable for use asa haemostatic paste, that the paste results in efficient and fasthaemostasis and that the paste extrudes easily from the applicatordevice so that the surgeon applying the paste can administer the pasteprecisely to a site of bleeding without using excessive force.

The paste of the present disclosure is superior to currently availablepastes such as Floseal and Surgiflo due to the fact that the paste ofthe present disclosure may be prepared simply by adding an amount of anaqueous medium to the dry composition, whereupon a ready-to-usehaemostatic paste forms spontaneously, i.e. within less than about 30seconds, preferably within less than about 20 seconds, more preferredwithin less than about 10 seconds, even more preferred within less thanabout 5 seconds.

The quantity of liquid to be added to the dry composition may beadjusted by the skilled person. The paste so formed always has anoptimal consistency when the correct amount of liquid is added. This isnot always the case with the conventional pastes, where the consistencyof the paste may depend on the force applied and time spent mixing. Thatno mechanical mixing is required also means that less time is spentpreparing the paste, which in turn leads to increased patient safety,both due to the fact that the haemostatic paste can be applied to thepatient faster and that the simple preparation method decreases thelikelihood of mistakes being made during the preparation of thehaemostatic paste.

When thrombin is comprised within the dry composition, the disclosurefurther has the advantage over conventional pastes in that it avoids thetime-consuming and error-prone thrombin dilution and addition stepsinvolved in current methods for making haemostatic pastes.

In one embodiment the present disclosure relates to a method forarresting bleeding/promoting haemostasis in an individual in needthereof by application of the dry composition or the reconstituted pasteas disclosed herein to a site of bleeding.

The paste of the present disclosure may be used for any type of surgeryincluding general surgery, cardiothoracic surgery, vascular surgery,plastic surgery, paediatric surgery, colorectal surgery, transplantsurgery, surgical oncology, trauma surgery, endocrine surgery, breastsurgery, skin surgery, otolaryngology, gynaecology, oral andmaxillofacial surgery, dental Surgery, orthopaedic surgery,neurosurgery, ophthalmology, podiatric surgery, urology.

In one embodiment the present disclosure relates to a method forpromoting wound healing in an individual in need thereof by applicationof the dry composition or paste of the present disclosure to the wound.

A “wound” refers broadly to injuries to the skin and/or underlying(subcutaneous) tissue initiated in different ways (e.g., pressure soresfrom extended bed rest and wounds induced by trauma) and with varyingcharacteristics. Wounds may be classified into one of four gradesdepending on the depth of the wound: i) Grade I: wounds limited to theepithelium; ii) Grade II: wounds extending into the dermis; iii) GradeIII: wounds extending into the subcutaneous tissue; and iv) Grade IV (orfull-thickness wounds): wounds wherein bones are exposed (e.g., a bonypressure point such as the greater trochanter or the sacrum). Thepresent disclosure relates to treatment of any type of wound mentionedabove using the dry composition or reconstituted paste of the presentinvention.

The treatment of a wound can in principle result in healing of the woundor in accelerated healing of the wound. The accelerated healing can be aresult of e.g. administration of a wound-healing promoting substance.Alternatively, the wound healing can be promoted by preventing bacterialor viral infection, or by reducing the risk of such an infection whichwould otherwise have prolonged the wound treatment process.

In one embodiment the present disclosure relates to a method forpromoting bone and/or tendon and/or tissue healing in an individual inneed thereof by application of the dry composition or paste of thepresent disclosure to the injured bone, tendon or tissue.

The “individual” referred to herein may be any mammal, including, butnot limited to, mammals of the order Rodentia, such as mice andhamsters, and mammals of the order Logomorpha, such as rabbits. It ispreferred that the mammals are from the order Carnivora, includingFelines (cats) and Canines (dogs). It is more preferred that the mammalsare from the order Artiodactyla, including Bovines (cows) and Swines(pigs) or of the order Perssodactyla, including Equines (horses). It ismost preferred that the mammals are of the order Primates, Ceboids, orSimoids (monkeys) or of the order Anthropoids (humans and apes). Anespecially preferred mammal is the human.

Kit

The present disclosure further relates to a kit comprising the drycomposition and an amount of aqueous medium matched to the amount of thedry composition so that upon addition of the aqueous medium, a paste ofa suitable consistency will form spontaneously, i.e. within seconds.Preferably, the kit is for use in the promotion of haemostasis or foruse in wound, bone, tendon and/or tissue healing in an individual inneed thereof.

Hence, in one embodiment the present disclosure relates to a kitcomprising:

-   -   a) a container comprising the dry composition obtained by the        method of the present disclosure,    -   b) a container comprising an aqueous medium, and    -   c) optionally an outer packaging.

The aqueous medium used to reconstitute the paste may be e.g. beselected from water, saline, a CaCl₂ solution or a buffered aqueoussolution.

In one embodiment, the aqueous medium used to reconstitute the drycomposition is water. In one embodiment, the tonicity of the aqueousmedium is selected so that an essentially isotonic paste will form uponaddition of the aqueous medium to the dry composition. The tonicity ofthe paste is usually selected so that it is compatible with surgicaluse.

In one embodiment, the aqueous medium used to reconstitute the drycomposition is saline.

In one embodiment, the dry composition comprises thrombin.

In one embodiment, the aqueous medium used to reconstitute the drycomposition comprises an extrusion enhancer, such as albumin. In thiscase, the dry composition preferably does not contain an extrusionenhancer.

Items

-   1. A method of preparing a dry composition suitable for use in    haemostasis and wound healing comprising the sequential steps of:    -   a. providing a biocompatible polymer in powder form, one or more        polyols, an extrusion enhancer and an aqueous medium,    -   b. mixing the biocompatible polymer, the one or more polyols,        the extrusion enhancer and the aqueous medium to obtain a paste,        and    -   c. drying the paste.-   2. The method according to item 1, wherein the extrusion enhancer is    albumin, preferably human serum albumin.-   3. The method according to any of the preceding items, wherein the    paste prior to drying comprises the extrusion enhancer in an amount    of between about 0.1% w/w to about 10% w/w, preferably between about    0.5% w/w to about 5.0% w/w.-   4. The method according to any of the preceding items, wherein the    dry composition comprises from about 10% w/w to about 60% w/w of one    or more polyols, such as from about 20% w/w to about 50% w/w of one    or more polyols, for example from about 20% w/w to about 40% w/w,    such as from about 20% w/w to about 30% w/w of one or more polyols.-   5. The method according to any of the preceding items, wherein the    paste prior to drying comprises from about 3% w/w to about 20% w/w    of one or more polyols, such as from about 3% w/w to about 18% w/w    of one or more polyols, for example from about 3% w/w to about 17%    w/w of one or more polyols, such as from about 3% w/w to about 16%    w/w of one or more polyols, for example from about 3% w/w to about    15% w/w of one or more polyols.-   6. The method according to any of the preceding items, wherein the    biocompatible polymer is cross-linked.-   7. The method according to any of the preceding items, wherein the    biocompatible polymer is obtained from a cross-linked sponge.-   8. The method according to any of the preceding items, wherein the    biocompatible polymer is gelatine.-   9. The method according to any of the preceding items, wherein the    drying is freeze-drying.-   10. The method according to any of the preceding items, wherein the    paste prior to drying comprises:    -   a. from about 5% w/w to about 20% w/w of one or more polyols,    -   b. from about 0.5% w/w to about 5% w/w of an extrusion enhancer,        such as albumin,    -   c. from about 15% w/w to about 25% w/w of biocompatible polymer,        and    -   d. from about 60% w/w to about 80% w/w of water.-   11. The method according to any of the preceding items, wherein the    dry composition comprises less than about 5% w/w of water,    preferably less than about 3% w/w of water, more preferred less than    about 2% w/w of water, such as less than about 1% w/w of water.-   12. The method according to any of the preceding items, wherein the    one or more polyols is selected from sugar alcohols, sugars and/or    derivatives thereof.-   13. The method according to item 12, wherein the one or more sugar    alcohols is selected from the group consisting of glycol, glycerol,    erythritol, threitol, arabitol, xylitol, ribitol, mannitol,    sorbitol, dulcitol, fucitol, iditol, inositol, volemitol, isomalt,    maltitol, lactitol and polyglycitol.-   14. The method according to any of the preceding items, wherein the    one or more polyols is mannitol and optionally one or more further    polyols.-   15. The method according to any of the preceding items, wherein the    dry composition further comprises one or more bioactive agents that    stimulate haemostasis or wound, bone, tendon and/or tissue healing.-   16. The method according to item 15, wherein the bioactive agent is    thrombin.-   17. The method according to any of the preceding items, wherein the    aqueous medium is selected from the group consisting of water,    saline, a calcium chloride solution and a buffered aqueous medium.-   18. The method according to any of the preceding items, wherein the    method comprises a further step of placing the dry composition into    an outer packaging, such as an aluminium foil packaging.-   19. The method according to any of the preceding items, wherein the    method comprises a further step of sterilising the dry composition.-   20. The method according to any of the preceding items, wherein the    paste is filled into and dried within an applicator suitable for    dispensing compositions in paste form, such as a syringe.-   21. The method according to any of the preceding items, wherein the    dry composition reconstitutes to form a paste without mechanical    mixing within less than about 30 seconds, preferably within less    than about 20 seconds, more preferred within less than about 10    seconds, even more preferred within less than about 5 seconds.-   22. A dry composition obtained or obtainable by the method of any of    the preceding items.-   23. A dry composition comprising a biocompatible polymer, an    extrusion enhancer such as albumin and one or more polyols.-   24. The dry composition according to item 23, wherein the    composition comprises:    -   a. from about 10% w/w to about 60% w/w of one or more polyols,    -   b. from about 0.3% w/w to about 30% w/w of an extrusion enhancer        such as albumin, preferably about 2% w/w to about 15% w/w,    -   c. from about 40% w/w to about 80% w/w of a biocompatible        polymer, and    -   d. from about 0.1% w/w to about 5% w/w of water.-   25. The dry composition according to any of items 22 to 24, wherein    the dry composition is present in an applicator suitable for    dispensing compositions in paste form, such as a syringe.-   26. The dry composition according to any of items 22 to 24, wherein    the dry composition is in the form of a sheet.-   27. Use of the dry composition obtained by the method of any of    items 1 to 21 or the dry composition of any of items 22 to 25 in the    preparation of a paste.-   28. The dry composition obtained by the method of any of items 1 to    21 or the dry composition of any of items 22 to 26 for use in the    promotion of haemostasis and/or wound, bone, tendon and/or tissue    healing in an individual in need thereof.-   29. A kit comprising:    -   a. a container comprising the dry composition obtained by the        method of any of items 1 to 21 or the dry composition according        to any of items 22 to 25,    -   b. a container comprising an aqueous medium, and    -   c. optionally an outer packaging.-   30. A method for reconstituting a dry composition comprising the    steps of:    -   a. providing a dry composition comprising a biocompatible        polymer, one or more polyols, and optionally an extrusion        enhancer such as albumin,    -   b. adding an aqueous medium to the dry composition, wherein the        aqueous medium optionally comprises an extrusion enhancer such        as albumin, with the proviso that an extrusion enhancer is        present in the dry composition and/or in the aqueous medium.-   31. The method according to item 30, wherein the dry composition is    as defined in any of items 22 to 26.

EXAMPLES Example 1. Haemostatic Pastes Comprising Varying Amounts ofMannitol and Glycerol

Materials

50 g Gelatine powder (milled crosslinked gelatine sponges)

200 ml buffer

Polyols

50% Benzalkoniumchloride (BAC)

0.9% Saline solution

x and y g Mannitol and Glycerol according to the following plan:

Formulation X: Mannitol Y: Glycerol [#] [g] [g] 1 20 3 2 5 3 3 12.5 5 412.5 5 5 20 5 6 5 7 7 5 5 8 20 7 9 12.5 3 10 12.5 7 11 0 0EquipmentFreeze dryer: Leybold-Heraus, Lyovac GT2 or Christ Alpha 1-4 LSCMixer: Kenwood, Major KM616MethodBuffer Solution:Add 2.0 g±0.1 g BAC (50%) to a 250 mL blue cap bottleAdd 98.0 g±0.5 g saline solution to the BACMix for 2 minutes using magnetic stirring—this is the BAC stock solutionAdd 123.0 g±0.5 g glycerol to a 2000 mL measuring flaskAdd 10 g±0.5 g BAC stock solutionAdd saline to the 2000 mL markPlace a stopper in the flask and turn it upside down a few timesMix by magnetic stirring for 5±1 minutesPaste:

Dissolve×g polyol(s) in 200 ml buffer solution under stirring in themixer. Add 50 g gelatine powder and mix with the dissolved polyol(s)until a homogeneous paste is obtained, approximately 5 minutes. Mixingof the paste was performed at room temperature, approximately 20° C.

Freeze-Drying:

The resulting paste is filled into 10 ml single use plastic syringes(5.5 ml per syringe) and placed at −30° C. for minimum 4 h. The frozenpaste is transferred to the freeze-dryer and freeze dried until dry for15 h.

Reconstitution:

The dry composition is reconstituted by adding 8 ml of liquid to eachsyringe, i.e. the amount of water which was removed from the compositionduring the freeze drying process is added. No mechanical mixing orstirring was used. The water was simply added to the dry composition andthe composition left untouched until a paste was re-formed.

Results

The different formulations were tested for time to reconstitution, i.e.the time needed for a paste suitable for haemostatic purposes tospontaneously form without mechanical agitation of any sorts.*Triplicate determination

Time to Formulation X: Mannitol Y: Glycerol reconstitution [#] [g] [g][sec]* 1 20 3 4 2 5 3 15 3 12.5 5 8 4 12.5 5 8.5 5 20 5 5 6 5 7 10 7 5 520 8 20 7 4 9 12.5 3 5 10 12.5 7 4 11 0 0 55

Formulation 11 is a negative control. The consistency of the formulation11 paste was clearly inferior to the consistency of the pastescontaining mannitol and glycerol in varying amounts.

Formulation 5

Formulation 5 had a spontaneous reconstitution time of 5 seconds. Thecontents of formulation 5 are specified in the table below in the paste(wet) and the dried composition (dry) respectively.

Content wet Content dry Content wet Content dry Formulation 5 [g] [g][%] [%] Gelatine 50.00 50.00 18.18 56.65 Mannitol 20.00 20.00 7.27 22.66Glycerol 12.30 12.30 4.47 13.94 (buffer) Glycerol 5.00 5.00 1.82 5.67(added) BAC 0.01 0.01 0.00 0.01 NaCl 0.01 0.01 0.00 0.01 H₂0 187.68 0.9468.25 1.06 SUM 275.00 88.26 100 100

The total percentage of glycerol in formulation 5 in the paste was thus6.29% and in the dried composition 19.61%.

The total polyol concentration, i.e. mannitol and glycerol, in the pastewas 13.56% and after drying 42.27%.

The polyol:gelatine ratio in the dry composition was approximately0.75:1.

Example 2. Mannitol and Glycerol

A paste was made, dried and reconstituted according to the methoddescribed in Example 1. The contents of the paste are shown in the tablebelow.

Content wet Content dry Content wet Content dry [g] [g] [%] [%] Gelatine50.00 50.00 18.52 60.05 Mannitol 20.00 20.00 7.41 24.02 Glycerol 12.3012.30 4.56 14.77 (buffer) Glycerol 0 0 0 0 (added) BAC 0.01 0.01 0.000.01 NaCl 0.01 0.01 0.00 0.01 H₂0 187.68 0.94 69.51 1.13 SUM 270.0083.26 100 100

The spontaneous reconstitution time of the paste made according to thetable above was 6 seconds.

The total polyol concentration, i.e. mannitol and glycerol, in the pastewas 11.97% and after drying 38.79%.

The polyol:gelatine ratio in the dry composition was approximately0.65:1.

Example 3. Mannitol

A paste was made, dried and reconstituted according to the methoddescribed in Example 1 with the exception that water was used instead ofthe buffer solution of Example 1. The contents of the paste are shown inthe table below.

Content wet Content dry Content wet Content dry [g] [g] [%] [%] Gelatine50.00 50.00 18.52 70.42 Mannitol 20.00 20.00 7.41 28.17 H₂0 200.00 1.0074.07 1.41 SUM 270.00 71.00 100 100

The spontaneous reconstitution time of the paste made according to thetable above was 7 seconds.

The results of the present example show that a paste of a suitableconsistency for haemostatic purposes can be obtained from a freeze driedpaste comprising only gelatine, water and a single polyol, in this casemannitol.

The polyol:gelatine ratio in the dry composition was approximately0.4:1.

Example 4. Trehalose and Glycerol

A paste was made, dried and reconstituted according to the methoddescribed in Example 1. The contents of the paste are shown in the tablebelow.

Content wet Content dry Content wet Content dry [g] [g] [%] [%] Gelatine50.00 50.00 18.52 60.05 Trehalose 20.00 20.00 7.41 24.02 Glycerol 12.3012.30 4.56 14.77 (buffer) Glycerol 0 0 0 0 (added) BAC 0.01 0.01 0.000.01 NaCl 0.01 0.01 0.00 0.01 H₂0 187.68 0.94 69.51 1.13 SUM 270.0083.26 100 100

The spontaneous reconstitution time of the paste made according to thetable above was 8 seconds.

The total polyol concentration, i.e. trehalose and glycerol, in thepaste was 11.97% and after drying 38.79%.

The polyol:gelatine ratio in the dry composition was approximately0.65:1.

Example 5. Thrombin

Thrombin was included in the formulation 5 paste of Example 1 at atheoretical concentration of 2500 IU/product (8 ml). The paste was madeat room temperature (approximately 20° C.) and mixed as described inExample 1.

The resulting paste was dried by freeze-drying and reconstituted asdescribed in Example 1. The thrombin activity was measured in thereconstituted paste. The results are shown in the table below.

Thrombin Activity - Freeze-dried composition in syringe [IU/product]2519.60 2884.94 2796.71 Mean activity: 2733.75

No loss of thrombin activity was measured in the reconstituted paste.

The results further show that it is not strictly necessary to performthe mixing of the paste at low temperatures to avoid loss of thrombinactivity as no decrease in thrombin activity was found when mixing wasperformed at ambient temperatures.

Example 6. Different Polyols

Pastes comprising different polyols were made, dried and reconstitutedessentially as described in Example 1 with the exception that H₂O withBAC was used instead of the buffer of example 1. The contents of thepaste are shown in the tables below.

Content wet Content dry Content wet Content dry [g] [g] [W/W %] [W/W %]Gelatine 50.00 50.00 18.52 70.41 Mannitol 20.00 20.00 7.41 28.17 BAC0.01 0.01 0.00 0.01 H₂0 200 1.00 74.07 1.41 SUM 270.01 71.01 100 100

Content wet Content dry Content wet Content dry [g] [g] [%] [%] Gelatine50.00 50.00 18.52 70.41 Xylitol 20.00 20.00 7.41 28.17 BAC 0.01 0.010.00 0.01 H₂0 200 1.00 74.07 1.41 SUM 270.01 71.01 100 100

Content wet Content dry Content wet Content dry [g] [g] [%] [%] Gelatine50.00 50.00 18.52 70.41 Trehalose 20.00 20.00 7.41 28.17 BAC 0.01 0.010.00 0.01 H₂0 200 1.00 74.07 1.41 SUM 270.01 71.01 100 100

Content wet Content dry Content wet Content dry [g] [g] [%] [%] Gelatine50.00 50.00 18.52 70.41 Maltitol 20.00 20.00 7.41 28.17 BAC 0.01 0.010.00 0.01 H₂0 200 1.00 74.07 1.41 SUM 270.01 71.01 100 100

Content wet Content dry Content wet Content dry [g] [g] [%] [%] Gelatine50.00 50.00 18.52 70.41 Sorbitol 20.00 20.00 7.41 28.17 BAC 0.01 0.010.00 0.01 H₂0 200 1.00 74.07 1.41 SUM 270.01 71.01 100 100

The polyol:gelatine ratio in the dry compositions was approximately0.4:1.

The spontaneous reconstitution time of the pastes comprising differentpolyols made according to the tables above is shown in the table belowand in FIG. 2. The experiments were repeated 5 times for each polyol.

Reconstitution Time in Seconds:

Mannitol Xylitol Trehalose Maltitol Sorbitol 1 7 14 11 14 29 2 9 31 2814 28 3 9 20 16 23 29 4 10 30 29 16 35 5 9 31 23 22 32 Average 8.8 25.221.4 17.8 30.6 Std 1.1 7.8 7.8 4.4 2.9

The experiment shows that different kinds of polyols can be used formaking a freeze-dried gelatine paste that will reconstitutespontaneously upon addition of water. The reconstituted paste has aconsistency suitable for direct use as a haemostatic paste.

The invention claimed is:
 1. A dry composition suitable for use inhaemostasis and wound healing comprising a frozen paste that has beenfreeze-dried, said dry composition comprising a biocompatible polymer,an extrusion enhancer and one or more polyols, wherein the drycomposition is obtained by the method comprising sequentially: a)providing a cross-linked biocompatible polymer in powder form, one ormore polyols selected from sugar alcohols or sugars, an extrusionenhancer selected from albumin, phosphatidylcholine, phosphatidylserine,lecithin or soy bean oil, and an aqueous medium; b) mixing thebiocompatible polymer, the one or more polyols, the extrusion enhancerand the aqueous medium to obtain a paste; and c) freeze-drying thepaste, wherein the dry composition comprises from about 10% w/w to about60% w/w of the one or more polyols, and wherein upon addition of anaqueous medium, the dry composition reconstitutes to form a pastewithout mechanical mixing.
 2. The dry composition according to claim 1,wherein the extrusion enhancer is albumin.
 3. The dry compositionaccording to claim 1, wherein the dry composition comprises from about0.3% w/w to about 30% w/w of the extrusion enhancer.
 4. The drycomposition according to claim 1, wherein the biocompatible polymer isobtained from a cross-linked sponge.
 5. The dry composition according toclaim 1, wherein the biocompatible polymer is gelatine.
 6. The drycomposition according to claim 1, wherein the dry composition comprisesless than about 5% w/w of water.
 7. The dry composition according toclaim 1, wherein the one or more polyols is one or more sugar alcoholsselected from glycol, glycerol, erythritol, threitol, arabitol, xylitol,ribitol, mannitol, sorbitol, dulcitol, fucitol, iditol, inositol,volemitol, isomalt, maltitol, lactitol and polyglycitol.
 8. The drycomposition according to claim 1, wherein the one or more polyols ismannitol and optionally one or more further polyols.
 9. The drycomposition according to claim 1, wherein the dry composition furthercomprises one or more bioactive agents that stimulate hemostasis orwound, bone, tendon and/or tissue healing.
 10. The dry compositionaccording to claim 9, wherein the bioactive agent is thrombin.
 11. Thedry composition according to claim 1, wherein the aqueous medium isselected from water, saline, a calcium chloride solution or a bufferedaqueous medium.
 12. The dry composition according to claim 1, whereinthe dry composition is comprised in an outer packaging.
 13. The drycomposition according to claim 1, wherein the dry composition is in theform of a sheet.
 14. The dry composition according to claim 1, whereinthe dry composition is sterile.
 15. The dry composition according toclaim 1, wherein the dry composition reconstitutes to form a pastewithout mechanical mixing within less than about 30 seconds.
 16. The drycomposition according to claim 1, wherein the dry composition comprises:a) from about 10% w/w to about 60% w/w of the one or more polyols, b)from about 0.3% w/w to about 30% w/w of the extrusion enhancer, c) fromabout 40% w/w to about 80% w/w of the biocompatible polymer, and d) fromabout 0.1% w/w to about 5% w/w of water.
 17. A syringe or applicatorcomprising a dry composition comprising a frozen paste that has beenfreeze-dried, said dry composition comprising a biocompatible polymer,an extrusion enhancer and one or more polyols, wherein the drycomposition is obtained by the method comprising sequentially: a)providing a cross-linked biocompatible polymer in powder form, one ormore polyols selected from sugar alcohols or sugars, an extrusionenhancer selected from albumin, phosphatidylcholine, phosphatidylserine,lecithin or soy bean oil, and an aqueous medium; b) mixing thebiocompatible polymer, the one or more polyols, the extrusion enhancerand the aqueous medium to obtain a paste; and c) freeze-drying the pasteto produce a dry composition, wherein the dry composition comprises fromabout 10% w/w to about 60% w/w of the one or more polyols, and whereinupon addition of an aqueous medium, the dry composition reconstitutes toform a paste without mechanical mixing.
 18. The syringe or applicatoraccording to claim 17, wherein the syringe or applicator is suitable fordispensing flowable haemostatic compositions.
 19. A haemostatic kit,comprising: a) a container comprising a dry composition comprising afrozen paste that has been freeze-dried, said dry composition comprisinga biocompatible polymer, an extrusion enhancer and from about 10% w/w toabout 60% w/w of one or more polyols, wherein the dry composition isobtained by the method comprising sequentially: i) providing across-linked biocompatible polymer in powder form, one or more polyolsselected from sugar alcohols or sugars, an extrusion enhancer selectedfrom albumin, phosphatidylcholine, phosphatidylserine, lecithin or soybean oil, and an aqueous medium; ii) mixing the biocompatible polymer,the one or more polyols, the extrusion enhancer and the aqueous mediumto obtain a paste; and iii) freeze-drying the paste to produce a drycomposition, wherein upon addition of an aqueous medium, the drycomposition reconstitutes to form a paste without mechanical mixing, b)a container comprising an aqueous medium, and c) optionally an outerpackaging.
 20. A dry composition suitable for use in haemostasis andwound healing comprising a frozen paste that has been freeze-dried, saiddry composition comprising: a) a cross-linked biocompatible polymer, b)an extrusion enhancer selected from albumin, phosphatidylcholine,phosphatidylserine, lecithin or soy bean oil, and c) from about 10% w/wto about 60% w/w of one or more polyols selected from sugar alcohols orsugars, wherein upon addition of an aqueous medium, the dry compositionreconstitutes to form a paste without mechanical mixing.
 21. A syringeor applicator comprising a dry composition comprising a frozen pastethat has been freeze-dried, said dry composition comprising: a) across-linked biocompatible polymer, b) an extrusion enhancer selectedfrom albumin, phosphatidylcholine, phosphatidylserine, lecithin or soybean oil, and c) from about 10% w/w to about 60% w/w of one or morepolyols selected from sugar alcohols or sugars, wherein upon addition ofan aqueous medium, the dry composition reconstitutes to form a pastewithout mechanical mixing.
 22. A haemostatic kit, comprising: a) acontainer comprising a dry composition comprising a frozen paste thathas been freeze-dried, said dry composition comprising: i) across-linked biocompatible polymer, ii) an extrusion enhancer selectedfrom albumin, phosphatidylcholine, phosphatidylserine, lecithin or soybean oil, and iii) from about 10% w/w to about 60% w/w of one or morepolyols selected from sugar alcohols or sugars, b) a containercomprising an aqueous medium, and c) optionally an outer packaging,wherein upon addition of an aqueous medium, the dry compositionreconstitutes to form a paste without mechanical mixing.